The cytochrome P-450-containing monoxygenase enzyme system is known to play key roles in the metabolism of drugs and the activation of carcinogens. This project is a continuation of our current studies on this enzyme system. The incorporation of purified cytochrome P-450 and reductase into the microsomal or other membranes will be studied to elucidate (1) the mechanism of the incorporation, (2) the nature of the binding, (3) the organization and interaction of monoxygenase enzymes, and (4) the rate-limiting step of drug oxidations. The enzyme topography of the microsomes will be studied with cross-linking techniques and a combined incorporation and cross-linking approach. The properties of the nuclear monoxygenase system will be characterized and the metabolism of benzo(a)pyrene by this enzyme system will be studied with high pressure liquid chromatography. The binding of the metabolically activated benzo(a)pyrene to nuclear DNA, RNA, histones, and nonhistone proteins will be studied using isolated rat liver nuclei. The effect of microsomes on the binding will be studied systematically to assess the roles of microsomes and nuclei in carcinogen activation. Similar approaches will also be used to study the sites of activation of aflatoxin B1.